DuPont B R, Grant S G, Oto S H, Bigbee W L, Jensen R H, Langlois R G
Biotechnology and Biomedical Research Division, Lawrence Livermore National Laboratory, Calif., USA.
Vox Sang. 1995;68(2):121-9. doi: 10.1111/j.1423-0410.1995.tb02563.x.
Glycophorin A (GPA) is an erythroid-lineage-specific membrane sialoglycoprotein which occurs in two allelic forms, M and N, which form the antigens of the MN blood group. Purified cDNAs and RNAs isolated from peripheral blood and erythroleukemia cell lines, HEL and K562, were used to develop an RT-PCR technique for amplifying GPA gene transcripts (GYPA). The relative expression of transcripts from the M and N alleles was determined using restriction analysis of these amplified products with four allele-specific restriction endonucleases. The use of this method permits the sensitive identification of GYPA transcripts in these cells and confirms GPA protein expression in the erythroleukemia cell lines and the MN phenotypes of individuals determined by immunolabeling with GPA allele-specific monoclonal antibodies. A novel restriction pattern was obtained using peripheral blood RNA from two individuals with a rare inherited variant allele, GPA Mg. Sequencing of the cDNA obtained using this method revealed a single C to A transversion in the fourth codon in the mature GYPA N coding sequence is responsible for the difference between GYPA Mg and GYPA N.
血型糖蛋白A（GPA）是一种红系特异性膜唾液糖蛋白，它以两种等位基因形式存在，即M和N，它们构成了MN血型系统的抗原。从外周血以及红白血病细胞系HEL和K562中分离得到的纯化cDNA和RNA，被用于开发一种逆转录聚合酶链反应（RT-PCR）技术，以扩增GPA基因转录本（GYPA）。使用四种等位基因特异性限制性内切酶对这些扩增产物进行限制性分析，从而确定M和N等位基因转录本的相对表达。该方法的使用能够灵敏地鉴定这些细胞中的GYPA转录本，并通过用GPA等位基因特异性单克隆抗体进行免疫标记，证实红白血病细胞系中GPA蛋白的表达以及个体的MN表型。使用来自两个具有罕见遗传变异等位基因GPA Mg的个体的外周血RNA，获得了一种新的限制性图谱。使用该方法获得的cDNA测序显示，成熟GYPA N编码序列的第四个密码子中发生了单个C到A的颠换，这导致了GYPA Mg和GYPA N之间的差异。
